Showing papers by "Victor Chang Cardiac Research Institute published in 1998"

Domain-specific gene disruption reveals critical regulation of neuregulin signaling by its cytoplasmic tail.

Abstract: Neuregulins are a multi-isoform family of growth factors that activate members of the erbB family of receptor tyrosine kinases. The membrane-anchored isoforms contain the receptor-activating ligand in their extracellular domain, a single membrane-spanning region, and a long cytoplasmic tail. To evaluate the potential biological role of the intracellular domain of the membrane-anchored neuregulin isoforms, we used a domain-specific gene disruption approach to produce a mouse line in which only the region of the neuregulin gene encoding almost the entire intracellular domain was disrupted. Consistent with previous reports in which all neuregulin isoforms were disrupted, the resulting homozygous neuregulin mutants died at E10.5 of circulatory failure and displayed defects in neural and cardiac development. To further understand these in vivo observations, we evaluated a similarly truncated neuregulin construct after transient expression in COS-7 cells. This cytoplasmic tail-deleted mutant, unlike wild-type neuregulin isoforms, was resistant to proteolytic release of its extracellular-domain ligand, a process required for erbB receptor activation. Thus, proteolytic processing of the membrane-bound neuregulin isoforms involved in cranial ganglia and heart embryogenesis is likely developmentally regulated and is critically controlled by their intracellular domain. This observation indicates that erbB receptor activation by membrane-bound neuregulins most likely involves a unique temporally and spatially regulated “inside-out” signaling process that is critical for processing and release of the extracellular-domain ligand.

Immunohistochemical localisation of α1B-adrenergic receptors in the rat iris

Abstract: Expression of the α1B-adrenergic receptor was investigated immunohistochemically in the rat iris, cornea and superior cervical ganglion by using antibodies raised in chickens immunised with a peptide corresponding to a portion of the 3rd intracellular loop common to the human, hamster and rat α1B-adrenergic receptor. Antibodies stained COS and HEK cell membranes of cells transfected with DNA encoding and expressing the hamster α1B-adrenergic receptor but not membranes from cells transfected with DNA encoding and expressing the rat α1A-adrenergic receptor or the rat α1D-adrenergic receptor. Staining was abolished by preincubation of the antibodies with the peptide used for immunisation. The distribution of α1B-adrenergic receptor was examined immunohistochemically with this antibody (1BI3) and a previously characterised antibody (Ab506) raised in rabbits against the carboxyl-terminal decapeptide of the receptor. In the iris, α1B-adrenergic receptor was detected in the dilator muscle, ciliary processes and posterior epithelium but no staining was observed in the superior cervical ganglion with either antibody. By contrast, differences in tissue staining between 1BI3 and Ab506 were observed for the sphincter muscle of the iris and for the cornea. 1BI3 stained both tissues intensely, whereas Ab506 only stained the cornea weakly and the sphincter not at all. Reverse transcription/polymerase chain reaction and nucleotide sequencing confirmed the presence of mRNA encoding the epitopes recognised by 1BI3 and Ab506 in cornea and other tissues. We conclude that (1) there is a good correlation between α1B-adrenergic receptor mRNA and protein expression in the iris, (2) mRNA, but not protein, is detected in the superior cervical ganglion and (3) additional processes may regulate receptor expression in the cornea.